Colon cancer formation and progression is the result of several molecular genetic alterations. Less important than the order of the accumulating genetic defects is the sum of those defects. Hyperplasia of the colon epithelium proceeds to adenoma, which will eventually develop to carcinoma and finally metastasize as genetic lesions accumulate. A number of genes involved in colorectal tumorigenesis have been delineated, either because they are known oncogenes (ras) or because they encode genes at a chromosomal locus known to be altered in colorectal tumors (p53, DCC, APC). Inherited mutations causing loss of function of some of these genes (p53, DCC, APC) greatly increase the risk of neoplasia in accordance with the "two-hit" hypothesis of Knudson. Sometimes, as with patients containing germline ablations of the APC gene, a clinically-observable disorder is recognized in which affected individuals are at extremely high risk for colon cancer at a relatively young age. However, most of the 130,000 cases of colon cancer that arise each year will do so without any such warning. Despite the slow progress of the disease, 65,000 people will die due to late presentation of symptoms. In order to better understand the molecular controls underlying colorectal cancer, and in order to offer a better chance for early detection of the disease, we are isolating and studying genes involved in colorectal tumorigenesis at various stages. Subtractive cDNA hybridization to enrich for populations of cDNAs expressed specifically in normal colon tissues, and specifically in tissues of the tumors, have been utilized. Unlike the methods used to identify the previously described genes, subtraction hybridization requires no a priori knowledge about the gene, its products, or its function. A number of genes whose possible role in colorectal cancer had not been suspected, but whose products were already known, have previously been identified. In this report, the characterization of novel genes which may play important roles very early in colorectal tumorigenesis is described.